Relationship Between TIGAR Transcript Level Jilld Meat Quality In Swine

Pork is clearly China’s main choice of meat, accounting for nearly three quarters of its overall meat consumption. Economic development and diversification, personalized consumption, increasing production and lean meat is unable to meet the growing consumer demand for quality and flavor of the meat, the focus is increasingly high. However, there exists in pursuit of raising production pork simply, while ignoring the animal health and welfare, which cause the animal highly sensitive to the ambient conditions. Therefore, leading the change of meat quality such as color, flavor, tenderness, juiciness, and the ratio of PSE meat increased. According to this, researching the meat quality mechanism and controlling and the prevention of poor quality meat, especially the PSE meat is very important.Most study shows that PSE meat is closely associated to the rapid post-mortem anaerobic glycolysis, so in present study, a newly discovered protein which can inhibit glycolysis-TIGAR was used to investigate the relationship between the transcription level of TIGAR mRNA and meat quality, from which to explore the inhibition to PSE meat. Meanwhile the prokaryotic expression for TIGAR was laid the groundwork for subsequent research. The main contents and results are as follows:（1） The P-actin mRNA as house-keeping gene was used to establish the relative quantification real-time reverse transcription PCR （also qPCR） method for TIGAR mRNA. According to pig TIGAR mRNA sequence （XM₀01926543） from GeneBank, a pair of primers was designed to amplify the TIGAR mRNA, then by pMD18-T vector, the TIGAR mRNA and β-actin mRNA standard substance were obtained. And by optimum conditions, the qPCR method for detection of porcine TIGAR mRNA was established. These results showed that the R2of standard curve of TIGAR and β-actin mRNA are0.9995and0.9996, and the amplification efficiency of the two gene were103.7%and104.1%, so the method established could use for detecting TIGAR mRNA transcript levels, and this also met the requirements of sample determination.（2） The developed method for detecting TIGAR mRNA was used to determinate the transcript levels of TIGAR mRNA at different slaughter time in Yimeng Black Swine and Durocxlandracex （Yorkshire）, and the meat quality indicators of the two breeds at different slaughter times were also determinated. Then the correlation between the TIGAR mRNA transcript levels and meat quality indicators were analyzed by SAS9.1. The results showed that TIGAR mRNA transcript levels and meat quality indicators had highly significant correlations （P<0.01） at different slaughter times. This indicated that the TIGAR mRNA transcript levels and the change of meat quality indicators have high degree of consistency.（3） The developed method for detecting TIGAR mRNA was used to detect5species TIGAR mRNA transcript levels and the meat quality indicators at45min after slaughter were also determinated. Then the correlation between the TIGAR mRNA transcript levels and meat quality indicators were analyzed by SAS9.1. The results showed that TIGAR mRNA transcript level is variants by breeds （P<0.05）, and higher transcript level and better meat quality had significant positive correlations. Local breeds such as Yimeng Black, Taihu had high TIGAR mRNA transcript levels and better meat quality, and it was less likely to produce PSE meat.（4） Prokaryotic expression for TIGAR protein. Codon optimized and synthetic TIGAR mRNA was cloned to pET-26b （+） expression vector, and then transformed into E. coli. BL21（DE3） to induce the TIGAR protein. The result of SDS-PAGE and Western-blot showed that the size of TIGAR protein was nearly30kD, and that was match to the expected size of the protein. These results indicated that the6xHis-TIGAR fusion proteins was obtained successfully, which lead to the groundwork for further study.